The activity of tyrosine hydroxylase, the rate-limiting enzyme in the biosynthesis of catecholamines, is dependent upon a reduced unconjugated pteridine, tetrahydrobiopterin. Utilizing rat brain and neuroblastoma preparations we have analyzed pteridine content and biosynthesis in order to elucidate the pathway by which these compounds are synthesized in neural tissues, with the eventual goal of understanding the cellular interactions between tyrosine hydroxylase and its pteridine cofactor.